Human interferon-alpha and -Beta (Hu-IFN-alpha, Hu-IFN-Beta) were among the first cytokines purified, cloned and introduced into the clinical arena. These interferons are in use for worldwide for use in treating various cancers, viral diseases, multiple sclerosis and other diseases. Knowledge of the receptor which binds these interferons and its signal transduction mechanisms are directly relevant to understanding the activities of the interferons. This proposal describes a procedure to isolate and identify all the constituents of the human Type I interferon (Hu-IFN-alpha/Beta) receptor. By a new technology, a yeast artificial chromosome (YAC) was isolated (alphaYAC) that expresses activity of a functional Type I human interferon receptor. The specific aims of this project are: to characterize the responses of the functional receptor to the Type I interferons; to determine the genes necessary for Type I interferon receptor function; to isolate the cDNA clones responsible for Type I IFN receptor activity; to reconstitute the receptor complex in heterologous cells (mouse and/or hamster cells); to carry out a structure/activity analysis of the receptor components by site-specific mutagenesis; to complement cells with mutations in the Type I interferon receptor and signal transduction pathways with the alphaYAC and cDNA clones; and to determine the expression of the various functional cDNA components in normal tissues and tumors. The alphaYAC clone provides the basis for these studies. Homologous recombination in yeast cells containing the alphaYAC will allow us to define the relevant genes responsible for Type I receptor function. It will be possible to reconstitute Type I human interferon receptor function by transfection of these cDNA expression vectors into heterologous cells (hamster and mouse cells); to delineate the various signal transduction mechanisms for the Type I interferons; and to evaluate their expression in normal and tumor tissues. Identification of all the components of the Type I interferon receptor will provide an understanding of the ligand-receptor interactions, the details of the signal transduction mechanisms and provide us with an opportunity to develop second generation interferons that exhibit fewer side effects and have improved efficacy.